Factor XIII supports platelet activation and enhances thrombus formation by matrix proteins under flow conditions.

BACKGROUND: Activated coagulation factor XIII (FXIIIa) is a transglutaminase that crosslinks fibrin at sites of vascular injury. FXIIIa also associates with blood platelets, although its role in platelet function is unclear and requires clarification. OBJECTIVES: To evaluate the ability of FXIIIa to support platelet adhesion and spreading under conditions of physiologic flow, and to identify the underpinning receptors and signaling events. METHODS AND RESULTS: Platelet adhesion to immobilized FXIIIa was measured by fluorescence microscopy, and signaling events were characterized by immunoblotting. Immobilized FXIIIa supported platelet adhesion and spreading under static conditions through mechanisms that were dually and differentially dependent on integrins α(IIb)ß(3) and α(v)ß(3). Platelet adhesion was independent of FXIIIa transglutaminase or protein disulfide isomerase activity. Moreover, adhesion was abolished by antibodies that prevented interaction with FXIIIa, but maintained when potential interactions with fibrinogen were blocked. Platelet adhesion to FXIIIa was reduced significantly by either the specific α(IIb)ß(3) antagonist tirofiban or the selective α(v)ß(3)-blocking antibody LM609, and abolished when they were used in combination. Importantly, platelet adhesion was preserved under venous and arterial flow conditions in which both integrins played essential roles. In contrast, FXIIIa stimulated the formation of filopodia and lamellipodia in adherent platelets that was mediated exclusively by α(IIb)ß(3) and eliminated by the Src-family inhibitor 4-amino-5-(4-methylphenyl-7-(t-butyl)pyrazolo(3,4-d)pyrimidine, indicating a tyrosine kinase-dependent mechanism. Crucially, under conditions of arterial shear, FXIIIa accentuated platelet recruitment by von Willebrand factor and collagen. CONCLUSIONS: Our data demonstrate a potential role for FXIIIa in supporting platelet adhesion at sites of vascular damage, particularly in association with other thrombogenic matrix proteins.

Bernard-Soulier syndrome in pregnancy.

Bernard-Soulier syndrome (BSS) is a rare autosomal recessively inherited bleeding disorder. Pregnancy in patients with BSS is characterized by ante-, intra-, or postpartum haemorrhage, which may be delayed and severe. There is no consensus in the management of BSS in pregnancy and so far only 16 pregnancies in nine patients have been described. We report a further three pregnancies in two women with the syndrome. We also outline our management of pregnant patients with BSS.

The rare coagulation disorders--review with guidelines for management from the United Kingdom Haemophilia Centre Doctors' Organisation.

The rare coagulation disorders are heritable abnormalities of haemostasis that may present significant difficulties in diagnosis and management. This review summarizes the current literature for disorders of fibrinogen, and deficiencies of prothrombin, factor V, FV + VIII, FVII, FX, the combined vitamin K-dependent factors, FXI and FXIII. Based on both collective clinical experience and the literature, guidelines for management of bleeding complications are suggested with specific advice for surgery, spontaneous bleeding, management of pregnancy and the neonate. We have chosen to include a section on Ehlers-Danlos Syndrome because haematologists may be consulted about bleeding manifestations in such patients.

Cutaneous B-cell lymphoma: pathological spectrum and clinical outcome in 51 consecutive patients.

The clinical behavior and optimal treatment of patients presenting with skin infiltration by B-cell lymphoma have not been established. To clarify this we assessed the clinical and laboratory features of 51 patients presenting with cutaneous infiltration by B-cell lymphoma. Follow-up data was available for 46 patients with a median age of 68 years (range 16-89 years) and a median follow-up of 32.5 months (range 5-123 months). Thirty-three of 51 (65%) patients had diffuse large B-cell lymphoma (DLBCL), and 15/51 (29%) had marginal zone lymphoma (MZL). The remaining 3 patients had follicular lymphoma, CLL, and post-transplant lymphoproliferative disease. Of the 33 patients with DLBCL, follow-up was available in 29; 24/29 (83%) had primary cutaneous disease, which was unifocal in 17/24 (71%). Following treatment, 8/24 (33%) of the primary cases relapsed. Of the 8 who relapsed, 7 had received local forms of treatment only. Follow-up data was available in 14/15 patients with MZL. 11/14 (79%) had primary cutaneous disease, which was unifocal in 8 (73%). Following treatment, 4 of these cases relapsed (36%); all had received local therapy only. It is evident from this study that a significant proportion ( reverse similar 20%) of patients who present with cutaneous infiltration by B-cell lymphoma have systemic disease. Staging is therefore mandatory in these patients. Approximately 1/3 patients with primary cutaneous DLBCL or MZL ultimately relapse, and relapse rates appear higher in those patients receiving local therapy only. Systemic or combined modality therapy may therefore be the most appropriate treatment at presentation. This should be assessed prospectively in randomized trials.

The development of targeted chemotherapy for CNS lymphoma-a pilot study of the IDARAM regimen.

PURPOSE: We have developed and evaluated a CNS-targeted chemotherapy regimen based on the pharmacokinetic properties of the individual drugs in the combination. PATIENTS AND METHODS: In a twin-track study, 16 patients with secondary CNS lymphoma (SCNSL) and 8 with primary CNS lymphoma (PCNSL) were treated with IDARAM which comprised idarubicin 10 mg/m(2) i.v., days 1 and 2; dexamethasone 100 mg, 12-h infusion, days 1, 2 and 3; cytosine arabinoside (ARA-C) 1.0 g/m(2), 1-h infusion, days 1 and 2; methotrexate 2.0 g/m(2), 6-h infusion, day 3 (with folinic acid rescue); and cytosine arabinoside 70 mg plus methotrexate 12 mg, intrathecally, days 1 and 8. Two cycles were delivered at 3-weekly intervals. After response assessment, patients received adjuvant cranial radiotherapy (40 Gy over 20 fractions). RESULTS: The series comprised 24 patients, 11 male and 13 female. Their median age was 53 years (range 21 to 73 years). Grade 4 neutropenia and thrombocytopenia occurred in the majority of patients treated. Of the eight PCNSL patients, seven achieved complete remission (CR). Four remained in CR at the time of this report with a median duration of follow-up of 25 months (range 11 to 42 months). Of the 16 SCNSL patients, 12 achieved CR. Seven patients remained in CR at the time of this report with a median duration of follow-up of 24 months (range 18 to 57 months). CONCLUSION: This study suggests that IDARAM is an effective regimen in both PCNSL and SCNSL and is suitable for further development and evaluation.

The impact of a 10-year audit cycle on blood usage in a district general hospital.

As clinical governance moves from concept to practice, it is emerging as a realistic strategy to promote and improve quality within the National Health Service, as well as satisfying the demand for external accountability. In the context of blood transfusion, the area of responsibility encompasses product liability, as well as efficient use of blood as a resource and transfusion as an appropriate clinical response. Clinical governance may be a modern catch phrase, but the principles it enshrines have long been established within blood transfusion, and in other aspects of haematology. Here, an audit cycle comprising four audits over a 10-year period to monitor the use of cross-matched blood in a large district general hospital is described. Initially, blood use was considered by hospital site, and by the surgical procedure for which it was requested. Later, the scope of the audit was expanded to consider usage by individual consultant. A standard of efficient use of cross-matched blood was taken to be a cross-match to transfusion ratio of < 1.5. The information was reviewed by the hospital transfusion committee, who have a key role in co-ordinating and assessing the practice of transfusion within a hospital. In this hospital, audit has been one of the main tools for improving practice, in particular by enabling the implementation and continuous revision of a maximum blood order schedule. Further, as the process of audit has developed, problem areas have been highlighted, and strategies to improve usage have been brought in with encouraging results. The audit is now being expanded again to include a greater focus on usage of cross-matched blood in the nonsurgical setting.

Waldenström macroglobulinemia. Development of diagnostic criteria and identification of prognostic factors.

To establish whether a combination of morphologic and immunophenotypic criteria could be developed to more precisely define Waldenström macroglobulinemia (WM) and prognostic factors, we retrospectively assessed the clinical and laboratory features of 111 cases of WM. Bone marrow infiltration by small lymphocytes was documented in each case; and diffuse, interstitial, nodular, and paratrabecular patterns of infiltration were documented in 58%, 32%, 6%, and 4% of cases, respectively. Ninety percent were characterized by a surface immunoglobulin-positive, CD19+CD20+CD5-CD10-CD23- immunophenotype. The median overall survival from diagnosis was 60 months; univariate analysis revealed the following adverse prognostic factors: older than 60 years, performance status more than 1, platelet count less than 100 x 10(3)/microL (< 100 x 10(9)/L), pancytopenia, and diffuse bone marrow infiltration. Associated median survival was 40, 38, 46, 28, and 59 months, respectively. Multivariate analysis revealed age, performance status, and platelet count as prognostically significant, but stratification of patients according to the International Prognostic Index had limited value. We suggest defining WM by the following criteria: IgM monoclonal gammopathy; bone marrow infiltration by small lymphocytes, plasmacytoid cells, and plasma cells in a diffuse, interstitial, or nodular pattern; and a surface immunoglobulin-positive, CD19+CD20+CD5-CD10-CD23- immunophenotype.

Autoimmune thrombocytopenia in Waldenström's macroglobulinemia.

Autoimmune phenomena are well-recognised complications of Waldenström's macroglobulinemia (WM) and IgM monoclonal gammopathy. Peripheral neuropathy and cold agglutinin hemolytic anemia are the most common reported and occur in 5-10% of patients. Autoimmune thrombocytopenia has been rarely reported in WM and its incidence is not known. In this study we report the case of a 67-year-old man who presented with autoimmune thrombocytopenia who was subsequently found to have WM. Laboratory investigation demonstrated that platelet-associated IgM (PAIgM) but not PAIgG was clearly elevated compared to normal controls. In addition the patient's serum reacted strongly with a panel of donor platelets analysed with an indirect platelet immunofluorescence assay utilising an anti-IgM secondary antibody. Glycoprotein specificity could not however be demonstrated by ELISA techniques for platelet glycoproteins IIbIIIa, IaIIa, IbIXa, and IV. We also reviewed the case records of 104 additional cases of WM diagnosed at our institution between 5/93 and 5/99. Three further cases with clinically significant autoimmune thrombocytopenia were identified. The overall incidence of autoimmune thrombocytopenia (4/105, 3.8%) in this cohort of patients was similar to the incidence of peripheral neuropathy (7/105, 6.7%) and cold agglutinins (3/105, 2.9%).

Molecular analysis of the genotype-phenotype relationship in factor VII deficiency.

Factor VII (FVII) deficiency is a rare haemorrhagic condition, normally inherited as an autosomal recessive trait, in which clinical presentation is highly variable and correlates poorly with laboratory phenotype. The FVII (F7) gene was sequenced in 48 unrelated individuals with FVII deficiency, yielding a total of 23 novel lesions including 15 missense mutations, 2 micro-deletions, 5 splice junction mutations and a single base-pair substitution in the 5' untranslated region. Family studies were performed in order to distinguish the contributions of individual mutant F7 alleles to the clinical and laboratory phenotypes. Specific missense mutations were evaluated by molecular modelling in the context of the FVIIa-tissue factor crystal structure. Single base-pair substitutions in splice sites and the 5' untranslated region were studied by in vitro splicing assay and luciferase reporter gene assay, respectively. All probands were also typed for four previously reported F7 polymorphisms. In the majority of cases of FVII deficiency studied here, consideration of both mutational and polymorphism data permitted the derivation of plausible explanations for the FVII activity and antigen levels measured in the laboratory. Inter-familial variation in FVII activity and the antigen levels of heterozygous relatives of probands was found to be significantly higher than intra-familial variation, consistent with the view that the nature of the F7 gene lesion(s) segregating in a given family is a prime determinant of laboratory phenotype. Although no relationship could be discerned between laboratory phenotype and polymorphism genotype, the frequencies of the A2 and M2 polymorphic alleles were significantly higher in the FVII-deficient individuals tested than in controls. This suggests that the presence of these alleles may have served to increase the likelihood of pathological F7 gene lesions coming to clinical attention.

Incidence of iron-deficiency anaemia in infants in a prospective study in Jordan.

A high prevalence of iron-deficiency anaemia has been reported in Jordanian infants. A prospective study of infants in downtown Amman examined the relationship between anaemia in pregnancy and iron deficiency in infancy. The iron status of infants born to 107 anaemic (Hb < 11 g/dl) and 125 non-anaemic mothers was reviewed at 3, 6, 9 and 12 months. Indicators to define iron-deficiency anaemia were Hb < 11 g/dl and either plasma ferritin < 12 microg/l or zinc protoporphyrin (ZPP) > 35 microg/dl whole blood. Haemoglobin electrophoresis excluded haemoglobinopathy. There was 72% iron-deficiency anaemia throughout the year, significantly higher in infants born to anaemic mothers (81%; n = 91) compared with controls (65%; n = 112). At 12 months, 72% of the infants tested (n = 195) were anaemic. While 57% were identified as iron-deficient by research criteria of either ferritin or ZPP, only 37% were identified by ferritin alone, 40% by ZPP alone and 29% if both ferritin and ZPP were required to meet criteria. Most infant anaemia was identified as due to iron deficiency, supporting contextual setting as assisting diagnosis: infants in developing countries are recognised as vulnerable to iron deficiency. Using multiple criteria, more cases were identified when either ferritin or ZPP were abnormal than when one alone, or both parameters were required to meet research criteria.

Iron status, serum folate and B(12) values in pregnancy and postpartum: report from a study in Jordan.

BACKGROUND: A high prevalence of anemia has been reported in Jordan affecting women of childbearing age and infants/preschool children. This paper considers maternal iron, folate and B(12) status, with possible implications for both maternal and infant health. PATIENTS AND METHODS: A case-control study of infants born to anemic (Hb <11 g/dL) (n=107) and non-anemic mothers (n=125) from birth to one year, was conducted in a lower middle-class urban setting in Amman, Jordan. Maternal hematology included full blood count (FBC), plasma ferritin, transferrin saturation, serum folate and B(12) at term, and FBC and ferritin at 6 months postpartum. Serum B(12) was reassessed at 6 and 12 months postpartum if antenatal values were low. Infant FBC and ferritin were assessed in cord blood and at 3, 6, 9 and 12 months, and zinc protoporphyrin (ZPP) from 6 months. RESULTS: Anemic mothers (mean [SD] Hb 9.9 [0.7] g/dL) had significantly lower antenatal values for Hb, MCV, MCH, transferrin saturation, plasma ferritin and serum folate than non-anemic mothers (mean Hb 12.2 [0.9] g/dL), which persisted at 6 months postpartum. Antenatal B(12) values were low (<200 pg/mL) in 67% of samples (26% <100 pg/mL), evenly distributed between the groups, and, therefore, not related to maternal anemia. Low values persisted in 27% (n=127) and in 61% (n=31), respectively, at 6 and 12 months postpartum. Iron deficiency anemia (Hb <11 g/dL and either ferritin <12 mcg/L or ZPP >35 mcg/dL) affected 72% of infants, with significantly higher incidence in those born to anemic mothers. Ambiguous hematology in 11% of infants may have reflected other nutritional deficiencies, including vitamin B(12), where mothers had depleted values. CONCLUSION: Iron, folate and B(12) status should be monitored during pregnancy/lactation in developing countries, where nutritional deprivation is more prevalent and women of childbearing age often have a high fertility rate and inadequate inter-pregnancy interval to replenish body stores. Infant health may also be at risk through a compromised endowment of these micronutrients at birth.

Clinicopathological correlates of IgM paraproteinemias.

IgM paraproteinemia is considered to be the major defining feature of Waldenström's macroglobulinemia (WM), but it may also occur in other B-cell lymphoproliferative disorders. In this study we have reviewed the final pathological diagnosis of 106 patients with IgM paraproteinemia investigated in our laboratories between April 1993 and May 1999. In 22 of the 106 patients (20.8%), there was no clinical or laboratory evidence of an underlying lymphoproliferative disorder, and a diagnosis of monoclonal gammopathy of undetermined significance (MGUS) was therefore made. In 60 cases (56.6%), a diagnosis of WM was made, while in the remaining 24 patients, the final diagnosis was chronic lymphocytic leukemia (n = 10), diffuse large B-cell lymphoma (n = 5), extranodal marginal-zone lymphoma (n = 3), follicular lymphoma (n = 3), and mantle-cell lymphoma (n = 3). The median paraprotein concentration in patients with WM, MGUS, and "other" lymphoproliferative disorders was 13 g/L (range, 2-54), 6 g/L (range, 3-30), and 4.5 g/L (range, 3-61), respectively. It is clear that IgM paraproteins are demonstrable in all subtypes of peripheral B-cell disorders and, although paraprotein concentrations are generally higher in WM, there is considerable overlap. Immunophenotypic criteria are therefore essential for the accurate diagnosis of WM.

Eosinophilic myelodysplasia transforming to acute lymphoblastic leukaemia.

A 43 year old male presented with a marked eosinophilia and associated systemic symptoms. A diagnosis of myelodysplasia was made on the basis of bone marrow morphology and karyotype. Over a 12 month period the disease transformed into acute lymphoblastic leukaemia, confirmed by flow cytometry, cytochemistry, and immunohistochemistry. Karyotyping was abnormal with 5q- and -7 which persisted from diagnosis through to blastic transformation. He died following initial induction chemotherapy. Eosinophilic myelodysplasia is an uncommon condition in haematological practice and no previous report of lymphoblastic transformation has been found.

Anaemia during pregnancy as a risk factor for iron-deficiency anaemia in infancy: a case-control study in Jordan.

BACKGROUND: A high prevalence of 50-65% iron-deficiency anaemia in mothers and infants in Jordan was reported by the United Nations Relief and Works Agency (UNRWA) in 1990. Iron-deficiency in infancy has been shown to delay cognitive and psychomotor development with long-term consequences. While socioeconomic deprivation and inadequate nutrition are known underlying factors, it is unclear whether iron endowment at birth is compromised when mothers are anaemic, further jeopardizing iron status during infancy. A prospective case-control study of infants from birth to one year was conducted in a lower middle-class urban setting in Amman, Jordan. The study objective was to examine the relationship between maternal anaemia and iron-deficiency anaemia during infancy. METHOD: A sample of 107 anaemic (Hb < 11 g/dl) and 125 non-anaemic mothers was selected at 37 weeks' gestation and matched for age and parity, and infant data at birth obtained. The infants were reviewed at 3, 6, 9 and 12 months, to assess growth, current nutrition, infection rates and iron status. The main outcome measure was the incidence of iron-deficiency anaemia in the two groups of infants, defined in the study as Hb < 11 g/dl and either plasma ferritin < 12 mcg/l or zinc protoporphyrin > 35 mcg/dl. RESULTS: Iron endowment in cord blood samples appeared similar between the two groups. The incidence of iron-deficiency anaemia was very high in these infants, at 72% by research criteria, (51% if Hb < 10.5 g/dl), but significantly higher in the infants born to anaemic mothers at all stages of the year, with overall incidence of 81% (n = 91), compared to 65% in controls (n = 112). This was not explained by differences in environmental risk factors. Anaemic mothers had not recovered adequate iron status at 6 months' postpartum, with implications for future pregnancy iron demands. CONCLUSIONS: Anaemia during pregnancy compromises the health of mothers in traditional cultures, where women tend to have several children close together after marriage, with an inadequate interval to replenish nutritional stores. Their infants also appear to be at increased risk of developing iron-deficiency anaemia, undetected at birth.

Granulocytic sarcoma of the rectum: a rare complication of myelodysplasia.

A 67 year old man with myelodysplasia was admitted as an emergency with a six week history of rectal bleeding and diarrhoea. Barium enema showed an irregular polypoid filling defect in the lateral wall of the proximal rectum near the rectosigmoid junction. Histology showed this to be a granulocytic sarcoma (extramedullary granulocytic leukaemia; chloroma) infiltrating the bowel. A low index of suspicion of this lesion results in an incorrect diagnosis in many such cases. A chloroacetate esterase immunoperoxidase stain will confirm the granulocytic nature of the tumour cells.

Incidence of hereditary bleeding disorders in Bradford, UK: variation with ethnic group.

We have studied the number of patients registered with congenital bleeding disorders at the Haemophilia Centre, Bradford, UK, according to ethnic group. The large Pakistani population in Bradford presents a different spectrum of disorders compared with the indigenous Caucasian population with a significantly higher number of cases of factor VII deficiency and platelet disorders. Other haemophilia centres in the developed world serving large immigrant communities may also manage increased numbers of these rarer disorders with similar implications for resource allocation.